2,2,2-trifluoroacetic acid 1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene] hydrazide polymorphs and method of making the same

ABSTRACT

Crystalline polymorph forms of neurotrophic agent 2,2,2-trifluoroacetic acid 1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147), and process for producing the crystalline polymorphic form are provided.

RELATED PATENT APPLICATIONS

This patent application claims the benefit of, and is a U.S. nationalphase of, International PCT Patent Application No. PCT/US2019/018835filed Feb. 20, 2019, entitled 2,2,2-TRIFLUOROACETIC ACID1-(2,4-DIMETHYLPHENYL)-2-[(3-METHOXYPHENYL) METHYLENE] HYDRAZIDEPOLYMORPHS AND METHOD OF MAKING THE SAME; which claims the benefit ofU.S. Provisional Patent Application No. 62/633,441 filed on Feb. 21,2018, entitled 2,2,2-TRIFLUOROACETIC ACID1-(2,4-DIMETHYLPHENYL)-2-[(3-METHOXYPHENYL)METHYLENE] HYDRAZIDEPOLYMORPHS AND METHOD OF MAKING THE SAME. The entire content of theforegoing application is incorporated herein by reference, including alltext, tables and drawings.

BACKGROUND

The present disclosure relates to polymorph forms of a pharmaceuticalactive agent. In particular, the present disclosure relates to polymorphforms of neuroprotective agent 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene] hydrazide (J147).

2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene] hydrazide (J147)is a potent orally active neurotrophic agent discovered during screeningfor efficacy in cellular models of age-associated pathologies and has astructure given by Formula I:

J147 is broadly neuroprotective, and exhibited activity in assaysindicating distinct neurotoxicity pathways related to aging andneurodegenerative diseases, with EC₅₀ between 10 and 200 nM. It has beenindicated to improve memory in normal rodents, and prevent the loss ofsynaptic proteins and cognitive decline in a transgenic AD mouse model.Furthermore, it has displayed neuroprotective, neuroanti-inflammatory,and LTP-enhancing activity.

The neurotrophic and nootropic effects have been associated withincreases in BDNF levels and BDNF responsive proteins. Interestingly,despite this mechanism of action, J147's neuroprotective effects havebeen observed to be independent of TrkB receptor activation. J147 hasbeen indicated to reduce soluble Aβ40 and Aβ42 levels, and it iscurrently being researched for potential applications in treating ALS.

SUMMARY

The present disclosure is directed to a crystalline Form II of2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147),and methods of making thereof.

Certain embodiments of the present disclosure provide a method of makingcrystalline Form II of 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene] hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at13.37, 18.47, and 23.34+/−0.2 degrees 2-theta, the method comprising:providing a slurry comprising saturated amorphous or crystalline Form Iof 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene] hydrazide (J147)in a solvent/anti-solvent mixture; and mixing the slurry to providecrystalline Form II.

Certain embodiments of the present disclosure provide an isolatedcrystalline Form I of 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at11.85, 17.11, 17.79, and 23.40+/−0.2 degrees 2-theta.

BRIEF DESCRIPTION OF DRAWINGS

Various embodiments of the present disclosure will be described hereinbelow with reference to the figures wherein:

FIG. 1 shows the X-ray Diffraction (XRD) diffractogram of crystallineForm I of 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene] hydrazide (J147).

FIG. 2A shows a differential scanning calorimetry (DSC) thermogram ofcrystalline Form I of J147.

FIG. 2B shows a thermogravimetric (TGA) thermogram of crystalline Form Iof J147.

FIG. 3A shows dynamic vapor sorption isotherms of crystalline Form I ofJ147 using TGA.

FIG. 3B shows kinetic plots of crystalline Form I of J147 using TGA.

FIG. 4 shows the Fourier transform infrared (FTIR) spectrum ofcrystalline Form I of J147.

FIG. 5 shows the Raman spectrum of crystalline Form I of J147.

FIG. 6 shows proton nuclear magnetic resonance (NMR) spectrum ofcrystalline Form I of J147.

FIG. 7 shows graphical output using Debye cone integration using a twodimensional detector during XRD analysis of a sample of J147.

FIG. 8 shows a dendrogram of all XRD results in screening samples ofJ147.

FIG. 9 shows a cell plot of all XRD results in screening samples ofJ147.

FIG. 10 shows a cluster plot of all XRD results in screening samples ofJ147.

FIG. 11A shows a DSC thermogram of crystalline Form I of J147.

FIG. 11B shows the XRD spectrum of crystalline Form I J147.

FIG. 12A shows a representative photomicrograph of crystalline Form IJ147.

FIG. 12B shows another representative photomicrograph of crystallineForm I J147.

FIG. 13 shows the XRD spectrum of crystalline Form II J147.

FIG. 14A shows a DSC thermogram of crystalline Form II of J147.

FIG. 14B shows a TGA thermogram of crystalline Form II of J147.

FIG. 15 shows a representative photomicrograph of crystalline Form IIJ147.

FIG. 16A shows the moisture sorption-desorption isotherm of crystallineForm II J147 using dynamic vapor sorption (DVS) analysis.

FIG. 16B shows kinetic plots of crystalline Form II J147 using DVSanalysis.

FIG. 17 shows a computer predicted powder XRD pattern of a singlecrystal of Form II of J147. The predicted pattern matched the pattern ofexperimental pattern.

DETAILED DESCRIPTION

Embodiments herein are directed to various polymorph forms of2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147)and methods of preparing such forms. Discovery of polymorph forms ofactive pharmaceutical ingredients is recognized as an important practicein product development. Polymorph forms of J147 disclosed herein mayaffect various physicochemical properties of J147 including, withoutlimitation, hardness, stability, filterability, solubility,hygroscopicity, melting point, solid density and flowability. Alteringone or more physicochemical properties of J147 may lead to downstreamimprovements in both manufacture as well as having an impact onpharmacokinetics and other aspects of administering the API.

In some embodiments, there is provided an isolated crystalline Form I of2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at11.85, 17.11, 17.79, and 23.40+/−0.2 degrees 2-theta.

As used herein, “isolated” refers to the separation of a polymorph form,specifically of J147, to the substantial exclusion of other forms aswell as impurities and, if so desired, solvent. For example, an isolatedpolymorph may have a purity of at least about 95%, or about 98%, orabout 99%, or about 99.5%, including up to the limits of detection ofimpurities, or nominally 100% pure.

In some embodiments, isolated crystalline Form I may further compriseX-ray diffraction peaks located at 8.64, 13.36, 19.25, 21.64, and26.81+/−0.2 degrees 2-theta.

In some embodiments, there is provided isolated crystalline Form II of2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at13.37, 18.47, and 23.34+/−0.2 degrees 2-theta.

In some embodiments, isolated crystalline Form II may further compriseX-ray diffraction peaks located at 17.74, 20.39, 26.25, and 28.74+/−0.2degrees 2-theta.

Other XRD minor peaks in the spectra of Form I and Form II may bepresent, as disclosed herein below in the Examples.

In some embodiments, there are provided methods of making crystallineForm I of 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at11.85, 17.11, 17.79, and 23.40+/−0.2 degrees 2-theta, the methodcomprising recrystallizing 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide from anorganic solvent selected from the group consisting of nitromethane,methylethylketone, tetrahydrofuran, acetone, acetonitrile, heptane,isopropyl ether, isopropyl acetate, and chloroform.

In general, recrystallization techniques familiar to those skilled inthe art apply in the practice of the methods disclosed herein. Forexample, a sample of J147 may be dissolved in a minimal amount solventincluding dissolution at elevated temperatures for a given solvent. J147has been found to be reasonably soluble across an array of solventtypes, including polar protic and polar aprotic solvents. J147 generallyhas lower solubility in highly hydrophobic hydrocarbon solvents such asheptane, or at the opposite end, lower solubility in water. Thus, suchsolvents can serve as co-solvents or anti-solvents duringrecrystallization. Recrystallization may be performed with or withoutstirring, mixing, agitation, or the like.

In some embodiments, the methods of preparing Form I may further employwater as an anti-solvent. In some embodiments, a ratio of organicsolvent to anti-solvent water is in a range from about 4:1 to about 1:4.

In accordance with the methods disclosed herein and as exemplifiedbelow, a yield of Form I is obtainable in some embodiments in a rangefrom about 50% to about 100%, or about 90% to about 100%. In someembodiments, the yield may be at least about 95% based on the amount ofJ147 being recrystallized, or at least about 98%, or at least about 99%,or quantitative recovery.

In some embodiments, there are provided methods of making crystallineForm I of 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at11.85, 17.11, 17.79, and 23.40+/−0.2 degrees 2-theta, the methodcomprising recrystallizing 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2[(3-methoxyphenyl)methylene]hydrazide from asolvent-anti-solvent mixture comprising an alcohol as the solvent. Insome such embodiments, a ratio of solvent to the anti-solvent is in arange from about 4:1 to about 1:4. In some embodiments, the solventalcohol is a C₁-C₄ alcohol. Thus, for example, the alcohol may beselected from the group consisting of methanol, ethanol, 1-propanol,2-propanol, trifluoroethanol, 1-butanol and 2-butanol. In someembodiments the anti-solvent is water or heptane.

In accordance with the methods disclosed herein and as exemplifiedbelow, a yield of Form I is obtainable in some embodiments in a rangefrom about 50% to about 100%. In some embodiments, the yield may be atleast about 95% based on the amount of J147 being recrystallized, or atleast about 98%, or at least about 99%, or quantitative recovery.

In some embodiments, there are provided method of making crystallineForm II of 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at13.37, 18.47, and 23.34+/−0.2 degrees 2-theta, the method comprisingproviding a slurry comprising a saturated crystalline Form I of2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) ina solvent/anti-solvent mixture comprising water as anti-solvent andmixing the slurry at ambient temperature to provide crystalline Form II.

As used herein, “mixing” is broadly intended to include mixing,stirring, agitation and the like.

In some embodiments, Form II can be accessed via amorphous J147.

In some embodiments, the methods to access Form II of J147 may employ asolvent which is selected from the group consisting of an alcohol,dimethylformamide (DMF), and dimethylacetamide (DMA).

In some embodiments, the methods to access Form II of J147 involve theuse of a solvent alcohol which is selected from the group consisting ofmethanol, ethanol, trifluoroethanol, 1-propanol, and 2-propanol.

In some embodiments, the methods to access Form II of J147 a ratio ofthe solvent/anti-solvent mixture is in a range from about 1:2 to about1:1.

In some embodiments, the methods to access Form II of J147 includesmixing the slurry of saturated J147 over several days such as about 6days. In some embodiments, mixing may optionally be accompanied byheating. However, ambient laboratory conditions, i.e., about 25° C. aregenerally sufficient. In some embodiments, Form II may be accessible inreasonable purity and quantities after about 3 days, or about 4 days, orabout 5 days. Naturally, Form II may also be isolated at intervalslonger than 6 days if so desired, including about 7 days or about 8days.

In some embodiments, methods for forming Form II from the slurryincludes the use of an apparatus comprising a vessel connected to are-circulation system. In some embodiments, the recirculation can beconducted through a homogenization apparatus in which a shear force isapplied. The homogenization apparatus may comprise a stator and arotatable rotor, and the high-shear mixing force is applied by rotatingthe rotor at a speed of more than 250 rpm. The rotor can also be rotatedas speeds of more than 500 rpm and more than 1,000 rpm.

Re-circulating the slurry may comprise regulating the flow of slurrythrough the outlet and the inlet of the vessel. The energy for there-circulation can be provided by a pump. Conventional flow regulationmechanisms such as metering pumps, valves, and the like may be used forthis purpose. The process can also be conducted in a continuous mode.

In embodiments, Form I or amorphous J147 may be in a supersaturatedsolution dissolved in a solvent. This solution may be mixed with ananti-solvent solution. The anti-solvent refers to any solvent in whichthe chemical material has a poor solubility. It may be a mixture ofanti-solvents and solvents. For example, the anti-solvent may comprisewater or heptane. Mixing the solutions reduces the solubility of thematerial in the solvent mixture, causing it to crystallize out.

In embodiments, methods may also include the step of introducing seedcrystals into the vessel to facilitate crystallization. The seedcrystals may be placed into the supersaturated solution or into ananti-solvent. These seed crystals are selected to be insoluble in theindividual solvents and in the solvent mixture.

Mixing may comprise regulating the flow of the solution into the vessel.Conventional flow regulation mechanisms such as metering pumps, valves,and the like may be used for this purpose.

The temperature may be adjusted prior to their introduction into thevessel. This may be achieved by any conventional temperature adjustingequipment, such as a heater or a cooling bath associated with thesolution source.

In embodiments, the re-circulation system may comprise a homogenizationapparatus; outlet means for transferring the slurry from the vessel tothe homogenization apparatus; and inlet means for receiving the slurryfrom the homogenization apparatus into the vessel. The homogenizationapparatus may comprise a stator and a rotatable rotor, and means forapplying a high-shear mixing force by rotating the rotor. The high-shearmixing force can be applied by rotating the rotor at a speed of morethan 250 rpm. The rotor can also be rotated as speeds of more than 500rpm and more than 1,000 rpm.

The apparatus may include a means for regulating the flow of slurrythrough the homogenization apparatus. Conventional flow regulationmechanisms such as metering pumps, valves, and the like may be used forthis purpose. The apparatus may also include a means for adjusting thetemperature of the slurry in the vessel. This may be achieved by anyconventional temperature adjusting equipment, such as a heater or acooling bath associated with the solution source or the vessel.

In accordance with the methods disclosed herein and as exemplifiedbelow, in some embodiments, a yield of Form II is obtainable in a rangefrom about 50% to about 100%, or about 90% to about 100%. In someembodiments, the yield may be at least about 95% based on the amount ofJ147 being recrystallized, or at least about 98%, or at least about 99%,or quantitative recovery.

The following Examples are being submitted to illustrate embodiments ofthe present disclosure. These Examples are intended to be illustrativeonly and are not intended to limit the scope of the present disclosure.Also, parts and percentages are by weight unless otherwise indicated. Asused herein, “room temperature” refers to a temperature of from about20° C. to about 25° C.

EXAMPLES

This example describes the screening of J147 for polymorphic behavior.The screen was performed using solvent recrystallization, hydrationexperiments, competitive and non-competitive slurry experiments, andgrinding to manipulate the solid state form of the test material.Samples generated were characterized using differential scanningcalorimetry (DSC), polarized light microscopy, thermogravimetricanalysis (TGA), Fourier transform nuclear magnetic resonance (NMR)spectroscopy, powder X-ray diffraction (XRD), Fourier Transform Infraredspectroscopy (FTIR), Raman Spectroscopy and dynamic vaporsorption/desorption (DVS). The polymorph screen revealed that the J147is polymorphic, and several solid state forms were identified andcharacterized. The thermodynamically stable form was identified.

Experimental Methods

Microscopy: A Zeiss Universal and/or Olympus BX53 microscope configuredwith a polarized visible light source and polarizable analyzer were usedto evaluate the optical properties of the samples. The specimens weretypically mounted on a microscope slide with a cover glass. Observationsof particle/crystal size and shape and birefringence were recorded.

Hot Stage Microscopy (Hsm): A Linkam hot stage accessory was used intandem with the microscope. The specimens were mounted on a microscopeslide with a cover glass. The samples were heated from room temperaturethrough melting using a Linkam TMS 94 temperature control and Linksys 32data capture software system. Observations of possible phase change,melting, recrystallization, decomposition, etc. were recorded.

Proton Nuclear Magnetic Resonance Spectroscopy (1H-NMR): The sampleswere prepared by dissolving 1 to 10 mg of the API in deuteratedchloroform with 0.05% (v/v) tetramethylsilane (TMS). The spectra werecollected at ambient temperature on a Bruker 400 MHz NMR spectrometer.

Differential Scanning calorimetry (DSC): Differential Scanningcalorimetry (DSC) is a technique used to measure characteristic heatflux of a test article as it is scanned through a temperature gradientunder a controlled atmosphere. Thermal phase transitions such asendothermic melting and exothermic decomposition were recorded. The DSCdata were collected on a TA Instruments DSC. In general, samples in themass range of 1 to 10 mg were crimped in aluminum sample pans andscanned from 25 to approximately 150° C. at heating rates of 2, 10, 20,and 50° C./min using a nitrogen purge of 50 mL/min.

Thermogravimetric Analysis (TGA): Thermogravimetric Analysis involvesthe determination of the mass of a specimen as a function oftemperature. The TGA data were collected on a TA Instruments Q500 TGA.In general, samples in the mass range of 2 to 10 mg were placed in anopen, pre-tared platinum sample pan and attached by fine wire to amicrobalance. The sample was suspended in a furnace, which was heatedfrom 25 to about 250° C. at 10° C./min using a nitrogen purge at 100mL/min. The sample weight change as a function of temperature wasobserved.

X-Ray Powder Diffraction (XRD): X-ray diffraction is an analyticaltechnique used to study the crystalline nature of solid materials.X-rays incident on crystalline material are scattered in all directions.In certain directions, the scattered X-rays are constructivelyreinforced to form diffracted beams. The conditions for constructivediffraction are described by Bragg's Law and depend on the uniquecomposition and spatial arrangements of the crystal structure. As such,each molecular solid diffracts X-rays in different directions and atdifferent intensities resulting in a unique X-ray diffraction pattern. Avariable temperature hot stage was used to manipulate sample temperaturefor some experiments.

The X-ray powder diffraction patterns were obtained using a Bruker D8Discovery diffractometer equipped with an XYZ stage, laser videomicroscope for positioning, and a two-dimensional HiStar area Detectoror a scintillation detector. A Cu K a radiation 1.5406 angstrom sourceoperating at 40 kV and 40 mA was used to irradiate samples. The X-rayoptics consists of a Gobel mirror coupled with a pinhole collimator of0.5 mm Theta-theta continuous scans were employed with a sample-detectordistance of approximately 30 cm, which gives an effective range of 4-40.The samples were mounted in low background quartz plates.

Hygroscopicity-Dynamic Vapor Sorption (DVS): DVS is a gravimetricscreening technique that measures how quickly and how much of a solvent(water) is adsorbed by a sample. The relative humidity or vaporconcentration surrounding the sample is varied while the change in massof the sample is measured. A vapor sorption isotherm shows theequilibrium amount of vapor sorbed as a function of relativity humidity.The mass values at each relative humidity step are used to generate theisotherm. Isotherms are divided in two components: sorption forincreasing humidity steps and desorption for decreasing humidity steps.A plot of kinetic data is also supplied which shows the change in massand humidity as a function of time.

The samples were analyzed using a TA Q2000 automated dynamic vaporsorption analyzer. The samples were dried at 40° C. for 5 hours and thencooled to 25° C. with a dry nitrogen purge over them until they nolonger lost mass at 0% RH. The samples were then subjected to 0 to 95%RH, back to 0% RH at 25° C. in 5% RH steps.

Fourier Transform Infrared Spectroscopy (FTIR): The Infrared spectrawere obtained using a Nicolet 510 M-O Fourier transform infraredspectrometer, equipped with a Harrick Splitpea™ attenuated totalreflectance device. The spectra were acquired from 4000 to 400 cm⁻¹ witha resolution of 4 cm⁻¹; 128 scans were collected for each analysis.

Raman Spectroscopy: The Raman spectra were obtained with a Thermo DXRdispersive Raman spectrometer using laser excitation at 780 nm. Thespectra were acquired from 3300 to 100 cm⁻¹. The samples were analyzedas bulk powders.

Screening of J147 samples: Initial testing was performed by XRD, DSC,TGA, proton NMR, FTIR, and Raman spectrometry. X-ray powder diffractionwas used to examine the material to determine if it was crystalline.FIG. 1 shows the XRD pattern of this material which was crystalline anddesignated as Form I. The corresponding peaks and their abundance areshow in Table A below.

TABLE A Angle d value Intensity Intensity % 2-Theta ° Angstrom Cps %8.64 10.23 0.07 35.4 11.85 7.46 0.09 42.2 12.84 6.89 0.04 20.4 13.366.62 0.08 36.2 16.04 5.52 0.05 23.8 17.11 5.18 0.09 44.7 17.79 4.98 0.21100.0 19.25 4.61 0.08 38.4 21.64 4.10 0.08 37.6 23.40 3.80 0.09 42.026.81 3.32 0.07 35.3 29.76 3.00 0.06 28.7

The thermal behavior of Form I was determined by DSC and TGA. The DSCthermogram exhibited a melting endotherm with an onset of 62.6° C. Theendotherm was split with peak maxima at 66.5° C. and 74.7° C. The heatof fusion was 123.8 J/g. This split endotherm proved to be due to amixture of two polymorphic forms as observed by hot stage microscopy asdescribed further below. The scanning TGA thermogram indicated thesample was free of volatiles with a weight loss of less than 0.4 weight% from 25° C. to 104.7° C. FIGS. 2A and 2B show the DSC and TGAthermograms, respectively.

Dynamic vapor sorption isotherms and the kinetic plots are shown inFIGS. 3A and 3B, respectively. The material proved very hydrophobic anddid not appear to be prone to hydrate formation. A total weight loss ofapproximately 0.5% was observed at 95% RH. This unusual event (weightloss with high humidity) may be due to differences in the adsorptioncharacteristics of the sample and reference pans and not the J147sample.

The Fourier transform infrared (FTIR) spectrum is shown in FIG. 4. Basedon visual inspection the spectrum is consistent with structure. TheRaman spectrum is in agreement with the FTIR spectrum and is shown inFIG. 5. The proton NMR data is consistent with the structure of J147 andis shown in FIG. 6. The proton NMR data is also shown in tabulated formin Table B below.

TABLE B

Chemical Shift Peak Peak Area Tentative Proton (ppm) Type IntegralAssignment 7.298, 7.279 ~ 7.26 doublet doublet 4.94 a 7.256 singletchloroform ~7.26 to ~ 7.23 overlapping multiplets p, e, h ~ 7.22 to ~7.19 broad multiplet 1.02 m 7.134 to 7.110 multiplet 1.04 g 7.059, 7.039doublet 1.02 1 6.694 to 6.935 doublet doublet doublet 1.00 b 3.828singlet 3.06 d 2.419 singlet 3.05 o, r 2.085 singlet 3.08 1.547 broadpeak 0.58 water 1.416 broad peak 0.13 impurities 1.255 singlet 1.201.110 broad peak 0.38 0897 to 0.862 overlapping multiplets 0.17 0.000singlet NA TMS

Polymorph Screening: The screen was performed using solvent-basedrecrystallization followed by X-ray diffraction analysis of the solids.Suspension slurry experiments and grinding were also employed to searchfor additional solid state forms.

Visual Solubility Measurement: Approximately 80 mg of J147 was placedinto each of 25 vials. Solvent was added and vials were stirred forseveral minutes at ambient temperature, followed by visual observationfor remaining solids. The solvent was incrementally added until thesolids were dissolved, or until a maximum volume of solvent was addedand the experiment was terminated. These stock solutions were used toset up further panels of experiments. J147 was found to be quite solublein all solvents surveyed except for water. The visual solubility wasdetermined and shown in Table 1.

TABLE 1 Approximate Solvent Solubility mg/mL methanol >80 ethanol >80trifluoroethanol >80 1-propanol >80 2-propanol 40 1-butanol 40 2-butanol40 water <1.6 dimethyl formamide (DMF) >80 dimethylacetamide (DMA) >80butyl amine >80 pyridine >80 nitromethane >80 acetone >80 methyl ethylketone (MEK) >80 isopropyl ether >80 ethyl acetate >80 methyl tert butylether (MTBE) >80 isopropyl acetate >80 tetrahydrofuran (THF) >80acetonitrile (ACN) >80 dichloromethane (DCM) >80 chloroform >80toluene >80 heptane 8.0

Solvent Recrystallization: To perform the solvent-based portion of thepolymorph screen, the test material was recrystallized using varioussolvents under approximately 150 different crystal growth conditions.The scale of the recrystallization experiments was from approximately 6mL to 15 ml. The crystal growth conditions were changed by using binarygradient arrays of solvent mixtures. Saturated solutions were preparedby agitating excess (as possible) test material in contact with thevarious solvent systems at the saturation temperature. If solids did notcompletely dissolve in the solvent, the mother liquor was separated fromthe residual solids by filtration. The mother liquor was then heatedabove the saturation temperature (overheated) to dissolve any remainingsolids. The temperature of each solution was then adjusted to the growthtemperature and a controlled nitrogen shear flow was introduced to beginsolvent evaporation. Due to the high solubility of J147 in the majorityof solvents, an ambient growth temperature was used in all experiments.The recrystallization conditions for the six solvent based panels usedare summarized in Table 2. Each recrystallization panel contained from15 to 27 wells. The wells within each panel contained different solventcompositions. Because of the different solvent composition in each well,each well acted as a different crystal growth experiment. Based on theXRD analysis carried out, a new polymorph of J147 was discovered. Thefirst polymorph was designated as Form I while the second polymorph wasdesignated as Form II. Table 2 summarizes the recrystallization panelsfor solvent-based polymorph screening. The compositional solventmatrices for the six recrystallization panels used during thesolvent-based portion of the polymorph screen are shown in Tables 3 to8.

TABLE 2 No. Scale Saturation Overheat Growth N₂ Flow Panel of Wells (mL)Solvent Temp. (° C.) Temp. (° C.) Temp. (° C.) Rate (psi) 1 25 15 Single25 40 25 1 2 27 15 Binary 25 40 25 1 4 27  6 Binary 25 40 25 1 5 27  6Binary 25 40 25 1 6 15  6 Binary 25 40 25 1

TABLE 3 Well Solvent XRD Form Appearance 1 methanol amorphous orangeglass 2 ethanol amorphous clear glass 3 trifluoroethanol amorphous clearglass 4 1-propanol amorphous clear glass 5 2-propanol amorphous clearglass 6 1-butanol amorphous clear glass 7 2-butanol no sample no sample8 water no sample no sample 9 dimethyl formamide (DMF) amorphous clearglass 10 dimethylacetamide (DMA) amorphous clear glass 11 butyl amineamorphous orange glass 12 pyridine amorphous clear glass 13 nitromethaneI yellow solid 14 acetone amorphous clear glass 15 methyl ethyl ketone Iyellow solid (MEK) 16 isopropyl ether amorphous clear glass 17 ethylacetate amorphous clear glass 18 methyl tert butyl ether amorphous clearglass (MTBE) 19 isopropyl acetate amorphous clear glass 20tetrahydrofuran (THF) I white solid 21 acetonitrile (ACN) amorphousclear glass 22 dichloromethane (DCM) amorphous clear glass 23 chloroformI white solid 24 toluene amorphous clear glass 25 heptane amorphousclear glass

TABLE 4 Solvent Matrix and XRD Result for Recrystallization Panel 2Ratio of Solvents Solvent Sample ID 1 2 3 Co/Anti-Solvent methanol A12:3 7.5:7.5 3:12 water ethanol B 12:3 7.5:7.5 3:12 watertrifluoroethanol C 12:3 7.5:7.5 3:12 water 1-propanol D 12:3 7.5:7.53:12 water 2-propanol E 12:3 7.5:7.5 3:12 water 1-butanol F 12:3 7.5:7.53:12 water 2-butanol G 12:3 7.5:7.5 3:12 water DMF H 12:3 7.5:7.5 3:12water DMA I 12:3 7.5:7.5 3:12 water XRD Form Solvent Sample ID 1 2 3Co/Anti-Solvent methanol A I amorphous I water ethanol B I I I watertrifluoroethanol C amorphous I I water 1-propanol D I amorphous I water2-propanol E amorphous I I water 1-butanol F amorphous amorphousamorphous water 2-butanol G amorphous I amorphous water DMF H amorphousamorphous amorphous water DMA I amorphous amorphous amorphous water

TABLE 5 Solvent Matrix and XRD Result for Recrystallization Panel 3Ratio of Solvents Solvent Sample ID 1 2 3 Co/Anti-Solvent butyl amine A12:3 7.5:7.5 3:12 water pyridine B 12:3 7.5:7.5 3:12 water nitromethaneC 12:3 7.5:7.5 3:12 water acetone D 12:3 7.5:7.5 3:12 water isopropylacetate E 12:3 7.5:75 3:12 water THF F 12:3 7.5:7.5 3:12 water ACN G12:3 7.5:7.5 3:12 water heptane H 12:3 7.5:7.5 3:12 water isopropylether I 12:3 7.5:7.5 3:12 water XRD Form Solvent Sample ID 1 2 3Co/Anti-Solvent butyl amine A amorphous amorphous amorphous waterpyridine B amorphous amorphous amorphous water nitromethane C amorphousamorphous I water acetone D I amorphous amorphous water isopropylacetate E amorphous amorphous I water THF F amorphous amorphous I waterACN G amorphous amorphous I water heptane H I I amorphous waterisopropyl ether I I amorphous amorphous water

TABLE 6 Solvent Matrix and XRD Result for Recrystallization Panel 4Ratio of Solvents Solvent Sample ID 1 2 3 Co/Anti-Solvent methanol A12:3 7.5:7.5 3:12 heptane ethanol B 12:3 7.5:7.5 3:12 heptanetrifluoroethanol C 12:3 7.5:7.5 3:12 heptane 1-propanol D 12:3 75:7.53:12 heptane 2-propanol E 12:3 7.5:7.5 3:12 heptane 1-butanol F 12:37.5:7.5 3:12 heptane 2-butanol G 12:3 7.5:7.5 3:12 heptane DMF H 12:37.5:7.5 3:12 heptane DMA I 12:3 7.5:7.5 3:12 heptane XRD Form SolventSample ID 1 2 3 Co/Anti-Solvent methanol A amorphous amorphous amorphousheptane ethanol B amorphous amorphous amorphous heptane trifluoroethanolC amorphous I amorphous heptane 1-propanol D amorphous amorphousamorphous heptane 2-propanol E amorphous amorphous amorphous heptane1-butanol F amorphous amorphous not enough heptane material 2-butanol GI I not enough heptane material DMF H amorphous not enough amorphousheptane material DMA I amorphous amorphous amorphous heptane

TABLE 7 Solvent Matrix and XRD Result for Recrystallization Panel 5Ratio of Solvents Solvent Sample ID 1 2 3 Co/Anti-Solvent butyl amine A12:3 7.5:7.5 3:12 heptane pyridine B 12:3 7.5:7.5 3:12 heptanenitromethane C 12:3 7.5:7.5 3:12 heptane acetone D 12:3 7.5:7.5 3:12heptane MEK E 12:3 7.5:7.5 3:12 heptane isopropyl ether F 12:3 75:7.53:12 heptane ethyl acetate G 12:3 7.5:7.5 3:12 heptane MTBE H 12:37.5:7.5 3:12 heptane isopropyl acetate I 12:3 7.5:7.5 3:12 heptane XRDForm Solvent Sample ID 1 2 3 Co/Anti-Solvent butyl amine A amorphousamorphous amorphous heptane pyridine B amorphous amorphous not enoughheptane material nitromethane C amorphous amorphous not enough heptanematerial acetone D amorphous amorphous amorphous heptane MEK E amorphousamorphous amorphous heptane isopropyl ether F amorphous amorphousamorphous heptane ethyl acetate G amorphous amorphous amorphous heptaneMTBE H not enough amorphous amorphous heptane material isopropyl acetateI amorphous amorphous amorphous heptane

TABLE 8 Solvent Matrix and XRD Result for Recrystallization Panel 6Ratio of Solvents Solvent Sample ID 1 2 3 Co/Anti-Solvent THF A 12:37.5:7.5 3:12 heptane ACN B 12:3 7.5:7.5 3:12 heptane DCM C 12:3 7.5:7.53:12 heptane chloroform D 12:3 7.5:7.5 3:12 heptane toluene E 12:37.5:7.5 3:12 heptane XRD Form Solvent Sample ID 1 2 3 Co/Anti-SolventTHF A amorphous amorphous amorphous heptane ACN B amorphous amorphousamorphous heptane DCM C amorphous not enough amorphous heptane materialchloroform D not enough amorphous amorphous heptane material toluene Eamorphous amorphous amorphous heptane

Non-Competitive Slurry Experiments: In addition to the solventrecrystallization experiments, non-competitive slurry experiments wereperformed to search for new solid state forms. These experiments rely onsolubility differences of different polymorphic forms (if the compoundexists in different polymorphic forms). As such, only polymorphs havinga lower solubility (that is, are more stable) than the originalcrystalline form can result from a noncompetitive slurry experiment.

When a solid was mixed with solvent to create slurry, a saturatedsolution eventually resulted. The solution was saturated with respect tothe polymorphic form dissolved. However, the solution was supersaturatedwith respect to any polymorphic form that is more stable (more stableforms have lower solubility) than the polymorphic form initiallydissolved. Therefore, any of the more stable polymorphic forms cannucleate and precipitate from solution. In addition, noncompetitiveslurry experiments are often useful in identifying solvents that formsolvates with the compound.

The slurry experiments were performed by exposing excess suppliedmaterial to solvents and agitating the resulting suspensions for severaldays at ambient temperature. The solids were filtered (Whatman Grade 1,11 μm pore size) and analyzed by XRD to determine the resulting form(s).To avoid possible desolvation or physical change after isolation, thesamples were not dried before X-ray analysis. A summary ofnon-competitive slurry experiments is shown in Table 9.

TABLE 9 Vehicle Initial Form Duration Final Form Methanol/water (1:1) I6 days II Ethanol/water (1:1) I 6 days II Trifluoroethanol/water (1:1) I6 days II 1-propanol/water (1:1) I 6 days II 2-proponal/water (1:1) I 6days II DMF/water (1:1) I 6 days II DMA/water (1:1) I 6 days II

Based on their X-ray scattering behavior, all of the slurry experimentsof Form I resulted in Form II after 6 days of slurring. These dataindicate that Form II is more stable than Form I at ambient temperatureand pressure. No new polymorphs, solvates, or hydrates were isolated inthese experiments.

Solids generated from the solvent based recrystallization panels andsynthesized were analyzed by powder XRD. To mitigate preferred graineffects, a two dimensional detection system was used to collect all theXRD screening data. The two dimensional detector integrates along theconcentric Debye cones which helps reduce pattern variation. An exampleof the Debye cone integration using a two dimensional detector is shownin FIG. 7. If bright spots appear in the conical rings, it indicatesstrong preferred grain effects that can lead to considerable variabilityin the observed diffraction patterns including changes in peakintensities. Some samples of J147 exhibited preferred grain effectsbased on the appearance of the scattering behavior.

The results of this analysis revealed the material exists as twodifferent polymorphs. The polymorphs were designated as Forms I and II.The initially supplied material was designated as Form I. The resultingform designation for each individual (solvent-based) recrystallizationexperiment is shown in Tables 3 through 8 above. The XRD data collectedduring the study was evaluated using a full profile chemometrictreatment to determine if the crystalline form of the samples hadchanged during the experiment. To simplify the assessment, X-rayamorphous samples were not included in the chemometric treatment. Theanalysis entailed cluster analysis and multivariate statistics to grouptogether any patterns that were determined to be statistically the same.The results of this analysis are summarized in the dendrogram in FIG. 8,cell plot in FIG. 9, and cluster plot in FIG. 10. These figures providea pictorial cluster analysis of product similarity.

The chemometric analysis of the diffraction data categorized the samplesinto 3 different groups (or clusters) labeled A through C. A summaryshowing the number of members in each group (A through C) is shown inTable 10.

TABLE 10 Group Designated Form Members A II 30 B I 38 C Lowcrystallinity, 1 no form identified

The characteristics of each group/form are summarized as follows. Afterclassifying the data into different forms based on diffraction behavior,each form was studied to determine if other properties of the formscould be differentiated. The characterization of each form began bycomparing the diffraction data representative of each form with thatfrom the other forms. This was generally followed by DSC, TGA, DVSanalysis and microscopy.

Form I (Group B): The characteristic diffraction behavior of this formis shown in FIG. 1. The XRD patterns of the Form I samples were allcrystalline and very similar. This form was obtained from a variety ofsolvents in approximately 50% of the experiments. The DSC profile of theForm I samples exhibited a melting endotherm with an onset ofapproximately 63° C. In all but one of the Form I samples analyzed, theendotherm was split with peak maximums ranging from approximately 67° C.to 75° C. This split endotherm is believed to be due to a mixture of twopolymorphic forms as observed by hot stage microscopy. A representativeDSC thermogram is shown in FIG. 2A. A sample of Form I exhibited asingle endotherm in the DSC profile with an onset of melting atapproximately 63° C. The XRD pattern matched that of the other Form Isamples. FIGS. 11A and 11B show the DSC and XRD profiles, respectively,of this sample. The scanning TGA thermogram of Form I (FIG. 2B) showedit to be free from volatiles with a weight loss of less than 0.4 weight% from 25° C. to 104.7° C.

Microscopic evaluation of Form I showed a mixture of columnar and needleshaped particles ranging from approximately 10 to 300 microns in length.Upon heating this sample, the larger particles appear to melt atapproximately 67° C., with the smaller needles unmelted. FIGS. 12A and12B show representative photomicrographs at room temperature and 67° C.This along with the multiple endotherms in the DSC profile suggests themajority of the Form I samples may actually be a mixture of Forms I andII.

The dynamic vapor sorption isotherms and the kinetic plots for arepresentative sample of Form I are shown in FIGS. 3A and 3B,respectively. The material is very hydrophobic and does not appear to beprone to hydrate formation. A total weight loss of approximately 0.5%was observed at 95% RH. This unusual event (weight loss with highhumidity) is most likely due to differences in the adsorptioncharacteristics of the sample and reference pans and not the J147sample. A sample of Form I was placed in a 40° C. in oven over weekend.XRD data showed a conversion to Form II. A variable temperature (VT) XRDexperiment was performed on Form I. A sample of Form I was held in theXRD at 35° C. over a weekend with no form conversion observed. Overall,Form I was attributed to a dry crystalline, polymorphic form of thecompound.

Form II (Group A): The Form II polymorph was not obtained from therecrystallization screening experiments but was observed inapproximately 50% of all experiments. The XRD patterns representative ofForm II samples indicate that the samples were crystalline and verysimilar. FIG. 13 shows a characteristic XRD pattern of Form II. Thepeaks are summarized in Table C below.

TABLE C Angle d value Intensity Intensity % 2-Theta ° Angstrom Cps %12.00 7.37 0.08 6.9 13.37 6.62 0.52 47.4 17.74 5.00 0.14 13.1 18.47 4.800.19 17.4 20.39 4.35 0.18 16.7 23.34 3.81 1.10 100.0 26.25 3.39 0.1211.0 27.28 3.27 0.12 10.5 28.74 3.10 0.11 9.6 35.60 2.52 0.08 7.3 38.912.31 0.08 7.0

The DSC thermograms of this form exhibit a well-defined meltingendotherm with an extrapolated onset of approximately 74° C., a peakmaximum of approximately 75° C. and an enthalpy of fusion ofapproximately 68 J/g. FIG. 14A shows the DSC profile of Form II. The TGAthermogram for Form II showed it to be free from volatiles with a weightloss less than 0.1% from 25° C. to 75° C. FIG. 14B shows the TGAprofile. Microscopic evaluation of Form II showed birefringent needleshaped particles ranging from approximately 10 to 50 microns in length.FIG. 15 shows a representative photomicrograph of Form II crystals atroom temperature. Upon heating, one melting event was observed atapproximately 75° C. FIGS. 16A and 16B show the moisturesorption-desorption isotherm and the kinetic plots, respectively, forForm II. As seen with Form I, This form is also very hydrophobic anddoes not appear to be prone to hydrate formation A total weight loss ofapproximately 0.5% was observed at 95% RH. This unusual event (weightloss with high humidity) is most likely due to differences in theadsorption characteristics of the sample and reference pans and not theJ147 sample. The single crystal predicted powder XRD pattern matched thepattern of experimental Form II as shown in FIG. 17. A sample of Form IIwas placed in a 40° C. in oven over weekend. XRD data showed no formconversion. Overall, Form II was attributed to a dry crystallinepolymorphic form of the compound.

A number J147 samples were produced in this Example. The polymorphscreen recrystallization experiments produced either Form I or anamorphous form of J147 (as shown in Table 11). The Form II polymorph wasproduced by creating a saturated slurry of Form 1 and agitating/stirringfor several days at ambient temperature. Many of Form I samples alsocontained some amount of Form II indicating that the two polymorphs havea tendency to nucleate and grow concomitantly. Form II appears to be thethermodynamically stable form under ambient conditions based on theresults of the competitive slurry experiments.

TABLE 11 No. of Panel No. Experiments Form I Amorphous Panel 1 25 4 21Panel 2 27 12 15 Panel 3 27 8 19 Panel 4 27 3 24 Panel 5 27 None 26Panel 6 15 None 13 Total 148 27 118 % of total 100% 18% 80%

Competitive Slurry Experiments: In addition to the solventrecrystallization experiments, two competitive slurry experiments werealso performed to determine the most stable form. These experiments relyon the solubility differences of different polymorphic forms. As such,only polymorphic forms (and solvates) having a lower solubility (morestable) than the form initially dissolved can result from a competitiveslurry experiment.

The slurry experiments were performed by exposing excess material ofForms I and II to a small volume of solvent/water and agitating theresulting suspensions for several days at ambient temperature. Thesolids were filtered and analyzed by XRD and DSC to determine theresulting form. To avoid possible desolvation or physical change afterisolation, the sample was not dried before x-ray analysis. Table 12shows the results of the competitive slurry experiment.

TABLE 12 Initial Forms Slurry Final Form (XRD) Solvent Duration(XRD/DSC) I & II 1:1 ratio ethanol:water 4 days II I & II 1:1 ratio 8days II methanol:water

The competitive slurry experiments resulted in transformation of thesample to Form II. These results suggest that Form II is a less soluble,more thermodynamically stable polymorph relative to Form I at ambienttemperature and pressure.

Grinding: Form I was ground using a Crescent Wig-L-Bug ball mill for 1minute at 4800 oscillations per minute (3.2 m/s). XRD analysis showed notransformation under these conditions.

Recrystallization Using Heat: Amorphous/glass samples from Panel 1 wereplaced in a vacuum oven at 60° C. for 6 days. XRD analysis of thesesolids exhibited the XRD pattern of Form II. A sample of Form I wasplaced in a 40° C. in oven over a weekend. XRD data showed a conversionto Form II. A variable temperature (VT) XRD experiment was performed onForm I. A sample of Form I was held in the XRD at 35° C. over a weekendwith no form conversion observed.

Thermodynamic Relationship: DSC experiments were performed in order toobtain heat of fusion and melting data. This data can often be used todetermine if polymorphs exist in an enantiotropic or monotropicrelationship. The heat of fusion rule states that, if the higher meltingpolymorph has the lower heat of fusion, the two forms are enantiotropes.Conversely, if the higher melting polymorph has a higher heat of fusion,the two forms are monotropes. For a monotropic system, any transitionfrom one polymorph to another is irreversible. For an enantiotropicsystem, it may be possible to convert reversibly between the twopolymorphs on heating and cooling.

A sample of pure Form I and two samples of Form II were analyzed by DSCat a slow heating rate of 2° C./minute with similar sample sizes. Theaverage melting temperatures and heat of fusion data, from 10 runs, isshown in Table 13. These data indicate that Form I has a lower meltingtemperature and that Form II has a higher melting temperature. The heatof fusion values are very close with a high standard deviation for theForm I sample.

TABLE 13 Sample ID Onset (° C.) Heat of Fusion (J/g) Form I 63.7 ± 0.471.5 ± 7.6 Form II 73.9 ± 0.1 70.3 ± 0.9

Static Vapor Sorption Study: A dynamic vapor sorption study was done ina sealed humidity chamber with an automatic moisture sorption balance.Data collected during dynamic vapor sorption studies often are not atthermodynamic equilibrium. To determine if the Form I and Form IImaterial form a hydrate over time, samples were monitored in a 75%static humidity chamber.

In these studies, samples were stored in open Petri dishes in a chambercontaining a saturated salt solution to maintain the relative vaporpressure. A solution of saturated sodium chloride (75% RH) salt atambient temperature was used.

The samples were examined gravimetrically after 5 and 12 days. Afterboth time periods, neither Form I nor II showed significant weightchange, indicating no hydrate formation.

The raw diffraction data generated from the polymorph screeningexperiments were categorized into two polymorphic forms. Samples ofthese different forms were used to perform additional experiments (DSC,TGA, HSM, etc.) to refine the forms. A brief description of thediscovered polymorphic forms is summarized in Table 14.

TABLE 14 Form Designation Description Comments Form I Metastable PureForm I difficult to obtain. Tendency to nucleate and grow concomitantlywith Form II. Form II Thermodynamically Good crystallinity and thermalStable properties. Form II readily obtained from solvent slurry and heattreatment of Form I.

Table 14 summarizes the different solid state forms of J147 observedduring the study. The main result is the discovery of two anhydrouspolymorphic forms of J147. The polymorphic forms isolated in thisExample were designated I and II. At room temperature and pressure FormII is the thermodynamically form of J147. Form I is the metastable format room temperature and pressure.

Various experimental results have confirmed Form I is the metastableform at room temperature and pressure. Evidence of a transformation ofForm I to Form II was observed after approximately 3 days of storage at40° C. A competitive slurry of a 50/50 mix of Forms I and II in 2different solvent systems (at ambient temperature) showed conversion ofForm I to Form II after 4 and 8 days. Non-competitive slurry experimentsof Form I in 7 different solvents systems showed conversion to Form IIafter 6 days.

The screen entailed subjecting the material to solvent crystallization,heating, grinding, sorption experiments, and competitive andnoncompetitive slurry experiments. Overall, the J147 was recrystallizedunder more than 150 different crystal growth conditions and analyzedusing powder x-ray diffraction. The x-ray data was used to categorizethe samples into different groups. These groups were studied usingthermal, optical, spectroscopic, and other tools to elucidate the uniquesolid state forms of the API. In general, the J147 exhibits twodifferent polymorphic forms designated as Forms I and II in addition tothe amorphous form. No solvates or hydrates were uncovered in thisExample. Of the two polymorphic forms, the thermodynamically stablepolymorph under ambient conditions was Form II.

Example of Preparation of Form II of J147

Batch Process: About 100 kg of crude J147 from its synthetic preparationwas evaporated twice from about 80 kg of ethanol. The crude product wastaken up in about 48 kg of ethanol and the batch temperature wasadjusted to 28° C. About 37 kg of water was added gradually to thebatch. The batch was held at about 30° C. for about 1.7 hours. A sampleof the batch was pulled from the reactor and solids precipitated byaddition of 45 mL of water. The solids obtained were added back to thebatch as seed crystals and the mixture stirred for 40 minutes at 30° C.An additional about 34 kg of water was added. The batch was held atabout 18° C. for about 58 hours and then cooled to about 10° C. foranother about 5.5 hours. Analysis of the resultant solids indicated thepresence of Form I. Form I was converted to Form II by heating theslurry to about 45° C. for about 16 hours and then cooling back to about10° C. and holding the batch at this temperature for about 3 hours.about 17.7 kg of solid Form II of J147 were recovered by filtrationafter washing and drying.

What is claimed is:
 1. A method of making crystalline Form II of2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at13.37, 18.47, and 23.34+/−0.2 degrees 2-theta, the method comprising:(a) providing a slurry comprising saturated amorphous or crystallineForm I of 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene] hydrazide (J147)in a mixture comprising a solvent and an anti-solvent; and (b) mixingthe slurry to provide the crystalline Form II.
 2. The method of claim 1,wherein the slurry comprises saturated J147 in the crystalline Form I.3. The method of claim 1, wherein the mixing of (b) is carried out at atemperature in a range of about 25° C. to about 50° C.
 4. The method ofclaim 1, wherein the mixing of (b) is carried out at a temperature in arange of about 40° C. to about 50° C.
 5. The method of claim 1, whereinthe anti-solvent is water.
 6. The method of claim 1, wherein the solventis an alcohol.
 7. The method of claim 6, wherein the alcohol is selectedfrom the group consisting of methanol, ethanol, trifluoroethanol,1-propanol, and 2-propanol.
 8. The method of claim 6, wherein thealcohol is ethanol.
 9. The method of claim 1, wherein the solvent isdimethylformamide (DMF) or dimethylacetamide (DMA).
 10. The method ofclaim 1, wherein the solvent is ethanol and the anti-solvent is water.11. The method of claim 1, wherein a ratio of the solvent to theanti-solvent in the mixture is in a range from about 4:1 to about 1:4.12. The method of claim 11, wherein a ratio is about 1:2.
 13. The methodof claim 1, wherein the mixing of (b) occurs over about 6 hours to about6 days.
 14. The method of claim 1, wherein a yield of the crystallineForm II is in a range from about 50% to about 100%.
 15. The method ofclaim 1, wherein a purity of the crystalline Form II is at least 98%.16. The method of claim 1, wherein the crystalline Form I of J147 has apowder X-ray diffraction pattern comprising peaks located at 11.85,17.11, 17.79, and 23.40+/−0.2 degrees 2-theta, and is obtained by:recrystallizing 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide from anorganic solvent selected from the group consisting of nitromethane,methylethylketone, tetrahydrofuran, acetone, acetonitrile, heptane,isopropyl ether, isopropyl acetate, and chloroform, and optionally inthe presence of water as an anti-solvent.
 17. A method of makingcrystalline Form I of 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at11.85, 17.11, 17.79, and 23.40+/−0.2 degrees 2-theta, the methodcomprising: recrystallizing 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide from anorganic solvent selected from the group consisting of nitromethane,methylethylketone, tetrahydrofuran, acetone, acetonitrile, heptane,isopropyl ether, isopropyl acetate, and chloroform, and optionallyfurther comprising water as anti-solvent.
 18. The method of claim 17,wherein the recrystallizing is performed with water as the anti-solventand the ratio of organic solvent to anti-solvent is in a range fromabout 4:1 to about 1:4.
 19. A method of making crystalline Form I of2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at11.85, 17.11, 17.79, and 23.40+/−0.2 degrees 2-theta, the methodcomprising: recrystallizing 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide from amixture comprising a solvent and an anti-solvent, wherein the solvent isan alcohol, and the anti-solvent is water or heptane.
 20. Isolatedcrystalline Form I of 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at11.85, 17.11, 17.79, and 23.40+/−0.2 degrees 2-theta, and wherein thepowder X-ray diffraction pattern further comprises X-ray diffractionpeaks located at 8.64, 13.36, 19.25, 21.64, and 26.81+/−0.2 degrees2-theta.
 21. Isolated crystalline Form II of 2,2,2-trifluoroacetic acid1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147),having a powder X-ray diffraction pattern comprising peaks located at13.37, 18.47, and 23.34+/−0.2 degrees 2-theta, and wherein the powderX-ray diffraction pattern further comprises X-ray diffraction peakslocated at 17.74, 20.39, 26.25, 21.64, and 28.74+/−0.2 degrees 2-theta.